The importance of PUFAs is undisputed. For example, certain PUFAs are important biological components of healthy cells and are recognized as: “essential” fatty acids that cannot be synthesized de novo in mammals and instead must be obtained either in the diet or derived by further desaturation and elongation of linoleic acid (LA; 18:2 ω-6) or α-linolenic acid (ALA; 18:3 ω-3); constituents of plasma membranes of cells, where they may be found in such forms as phospholipids or triacylglycerols; necessary for proper development (particularly in the developing infant brain) and for tissue formation and repair; and, precursors to several biologically active eicosanoids of importance in mammals (e.g., prostacyclins, eicosanoids, leukotrienes, prostaglandins). Additionally, a high intake of long-chain ω-3 PUFAs produces cardiovascular protective effects (Dyerberg, J. et al., Amer. J. Clin. Nutr., 28:958-966 (1975); Dyerberg, J. et al., Lancet, 2(8081):117-119 (Jul. 15, 1978); Shimokawa, H., World Rev. Nutr. Diet, 88:100-108 (2001); von Schacky, C. and Dyerberg, J., World Rev. Nutr. Diet, 88:90-99 (2001)). Numerous other studies document wide-ranging health benefits conferred by administration of ω-3 and/or ω-6 PUFAs against a variety of symptoms and diseases (e.g., asthma, psoriasis, eczema, diabetes, cancer).
A variety of different hosts including plants, algae, fungi and yeast are being investigated as means for commercial PUFA production. Genetic engineering has demonstrated that the natural abilities of some hosts (even those natively limited to LA and ALA fatty acid production) can be substantially enhanced to produce high levels of e.g., γ-linolenic acid (GLA; 18:3 ω-6), dihomo-γ-linolenic acid (DGLA; 20:3 ω-6), arachidonic acid (ARA; 20:4 ω-6), eicosapentaenoic acid (EPA; 20:5 ω-3), docosapentaenoic acid (DPA; 22:5 ω-3) and docosahexaenoic acid (DHA; 22:6 ω-3).
Whether ω-3/ω-6 PUFA production is the result of natural abilities or recombinant technology, both strategies may require conversion of ω-6 PUFAs into their ω-3 counterparts. Specifically, a Δ15 desaturase is responsible for the conversion of LA to ALA, while a Δ17 desaturase is responsible for the conversion of ARA to EPA (although some Δ17 desaturases can also use DGLA) as a substrate to produce eicosatetraenoic acid (ETA; 20:4 ω-3)). Both of these enzymes have a role in the Δ6 desaturase/Δ6 elongase pathway (which is predominantly found in algae, mosses, fungi, nematodes and humans and which is characterized by the production of GLA and/or stearidonic acid (STA; 18:4 ω-3)) and the Δ9 elongase/Δ8 desaturase pathway (which operates in some organisms, such as euglenoid species and which is characterized by the production of eicosadienoic acid (EDA; 20:2 ω-6) and/or eicosatrienoic acid (ETrA; 20:3 ω-3)) (FIG. 1).
Because of the role Δ17 desaturase enzymes play in enabling the synthesis of ω-3 fatty acids, there has been considerable effort to identify and characterize these enzymes from various sources. However, only a few Δ17 desaturases are presently known and these have been isolated from only two different taxonomic genera. Specifically, Patent Publication No. US 2003/0190733 describes a Δ17 desaturase from Saprolegnia diclina (see also GenBank Accession No. AY373823). PCT Publication No. WO 2005/083053 describes a Phytophthora infestans “ω3 desaturase” (see also GenBank Accession No. CAJ30870), while PCT Publication No. WO 2006/100241 describes a Phytophthora sojae “ω3 desaturase”, both of which appear to function as Δ17 desaturases. Also, commonly owned, co-pending application having U.S. patent application Ser. No. 11/787,772 (filed Apr. 18, 2007) discloses nucleic acid and amino acid sequences for Δ17 desaturases from Phytophthora sojae and Phytophthora ramorum. Thus, there is need for the identification and isolation of additional genes encoding Δ17 desaturases that will be suitable for heterologous expression in a variety of host organisms for use in the production of ω-3 fatty acids.
Applicants have solved the stated problem by isolating the gene encoding Δ17 desaturase from the oomycete, Pythium aphanidermatum. 